Volume 41, Issue 6, Pages 974-982 (December 2004) Suppression of transforming growth factor-β results in upregulation of transcription of regeneration factors after chronic liver injury  Toru Nakamura, Takato Ueno, Masaharu Sakamoto, Ryuichiro Sakata, Takuji Torimura, Osamu Hashimoto, Hikaru Ueno, Michio Sata  Journal of Hepatology  Volume 41, Issue 6, Pages 974-982 (December 2004) DOI: 10.1016/j.jhep.2004.08.015 Copyright © 2004 European Association for the Study of the Liver Terms and Conditions

Fig. 1 Immunohistochemical analysis of hepatocyte proliferation (A and B) and apoptosis (C and D) in DMN-treated rat cirrhotic liver of AdLacZ-treated rats (A and C) and AdTβ-TR-treated rats (B and D) at 1 week after gene transfer. Cell proliferation was analyzed by immunohistochemistry using a monoclonal antibody against Ki-67 (A and B). The number of Ki-67-positive hepatocytes in the AdTβ-TR-treated rats was significantly higher than in AdLacZ-treated rats. Apoptotic cells were detected in situ by labeling fragmented DNA with the terminal deoxynucleotidyl transferase staining technique (TUNEL method) (C and D). The number of TUNEL-positive apoptotic hepatocytes in AdTβ-TR-treated rats was significantly lower than in AdLacZ-treated control rats. The number of TUNEL-positive apoptotic hepatocytes in DMN-rats injected with saline (data not shown) was similar to that in DMN-rats injected with AdLacZ (Original magnification ×200). Journal of Hepatology 2004 41, 974-982DOI: (10.1016/j.jhep.2004.08.015) Copyright © 2004 European Association for the Study of the Liver Terms and Conditions

Fig. 2 Quantitative analysis of proliferation (Ki-67) (A) and apoptosis (TUNEL) (B) indices in sections of regenerative DMN-treated rat cirrhotic livers in AdLacZ-treated and AdTβ-TR-treated rats at 1 week after gene transfer. The Ki-67 labeling index for hepatocytes in AdTβ-TR-treated rats was significantly higher than in AdLacZ-treated rats (P<0.01). In contrast, there was more than 18-fold decrease in the apoptosis index in AdTβ-TR-treated rats compared to AdLacZ-treated rats (P<0.01). The Ki-67 labeling index and apoptosis index in DMN-rats injected with saline (data not shown) were similar to those in DMN-rats injected with AdLacZ. Data are mean ±SEM. Journal of Hepatology 2004 41, 974-982DOI: (10.1016/j.jhep.2004.08.015) Copyright © 2004 European Association for the Study of the Liver Terms and Conditions

Fig. 3 Quantitative real-time PCR analysis of mRNA expression levels of regenerating growth factors after gene transfer in AdTβ-TR- (closed bars) and AdLacZ- (opened bars) treated rat liver tissues. Results are expressed in arbitrary units, and data are mean±SEM values of triplicate experiments with n=15 rats per time point. *P<0.05, **P<0.01 compared with before gene transfer (baseline value), #P<0.05, and ##P<0.01 compared with AdLacZ-treated rats per time point. Journal of Hepatology 2004 41, 974-982DOI: (10.1016/j.jhep.2004.08.015) Copyright © 2004 European Association for the Study of the Liver Terms and Conditions

Fig. 4 Quantitative real-time PCR analysis of IL-6 mRNA expression level after gene transfer in AdTβ-TR- (closed bars) and AdLacZ- (opened bars) treated rat liver tissues. Results are expressed in arbitrary units, and data are mean±SEM values of triplicate experiments with n=15 rats per time point. *P<0.05, **P<0.01. Journal of Hepatology 2004 41, 974-982DOI: (10.1016/j.jhep.2004.08.015) Copyright © 2004 European Association for the Study of the Liver Terms and Conditions

Fig. 5 Northern blot analysis of mRNA expression levels of EGF, TGF-α, HGF and IL-6 after gene transfer in AdTβ-TR- and AdLacZ-treated rat liver tissues. Non-treated control rats are designed as 0 day. (A) Representative set of autoradiograms of the Northern blot analyses. The same experiments were repeated more than 3 times with similar results. (B) Densitometric analyses of the autoradiograms of the Northern blot analyses. The mRNA levels were standardized with GAPDH. Results are expressed in arbitrary units, and data are mean±SEM values of triplicate experiments with n=3 rats per time point. *P<0.05, **P<0.01. Journal of Hepatology 2004 41, 974-982DOI: (10.1016/j.jhep.2004.08.015) Copyright © 2004 European Association for the Study of the Liver Terms and Conditions